dUTP used for PCR - (Aug/12/2001 )
I had used the dTTP to do the PCR before, but now I had changed to use the dUTP and the PCR product can not be generated. Does any one know how the PCR can work which the nucleotide used is dUTP.
You have to be sure that the thermostable polymerase that you're using will incorporate dUTP into DNA, some enzymes like Pfx polymerase will not. I would also suggest using double the concentrations of dUTP in your reactions since Taq tends to have a lower affinity for dUTP. Good luck.
I have the same problem ... I can't get PCR products for AT rich sequences using dUTP instead of dTTP
* lowering annealing temperature
* Increasing dUTP concentration (twice as high as normal dNTPs)
* Trying different enzymes
Does anybody knows about an enzyme (or something else) that may solve this PCR problem?